Apolipoprotein E (APOE) is a major cholesterol carrier that supports lipid transport and injury repair in the brain. In the CNS, APOE is mainly produced by astrocytes, and transports cholesterol to neurons via APOE receptors, which are members of the low-density lipoprotein receptor (LDLR). However, during stress or injury, neuronal production of APOE protein increases. APOE polymorphic alleles are the main genetic determinants of late-onset Alzheimer disease (LOAD) risk: individuals carrying the ?4 allele are at increased risk of LOAD compared to those carrying the more common ?3 allele, whereas the ?2 allele decreases risk. Presence of the APOE ?4 allele is also associated with increased risk for cerebral amyloid angiopathy and age- related cognitive decline during normal aging. Despite these findings, the mechanism by which APOE specifically influences AD pathophysiology is poorly understood. Even more importantly, why APOE ?4 is associated with LOAD in some people, but not in others remains unknown. Recently, we examined participant- derived olfactory cells of APOE4 carriers compared to non-carriers and observed differentially higher postmitotic (GAP43+)/mature (OMP+) densities, mitochondrial dysfunctions, cell deaths among aging cells of ?4 compared non-?4 isoforms. Importantly, while ?4 isoforms showed greater susceptibility to A? oligomer (ABO)-induced injury, we observed that individual differences in the expression of DNA pol? (DNA repair gene) moderated the impact of ABO on mitochondrial dysfunction, abnormal proteostasis and cell death. This is consistent with our previous publication (Misiak et. al. 2017) demonstrating that haploinsufficency of DNA pol? exacerbated mitochondrial dysfunction and cell death in AD mice. Therefore, we hypothesize that (1) APOE4 contributes to olfactory dysfunction in preclinical AD through aberrant neuronal differentiation, with resultant susceptibility to greater mitochondrial and synaptic dysfunctions, DNA damages and neuronal death, as the neurons advance in age and become exposed to increasing amount of toxins (e.g. A?); and (2) individual differences in proficiency for DNA repairs contribute to individual differences on these neuronal outcomes. We propose a 5-year prospective study to acquire olfactory cells through non-invasive sampling of 200 non-demented older (60-75 years; 100 ?4 and 100 non-?4) adults with well characterized olfactory functions to investigate molecular mechanisms underlying heterogeneity in severity of preclinical AD pathophysiologic markers. Molecular studies in this proposal are a multimodal combination of transcriptomic, RNA-based epigenomic, protein validation, mitochondrial profiling and enzymatic studies, aimed at uncovering novel preclinical mechanisms. In Aim 1, we will investigate molecular mechanisms underlying APOE isoform-specific effects on olfactory ? (a) differentiation (b) neurite morphology and axonal outgrowth (c) whole-cell reactive oxygen species (ROS) (d) mitochondrial function (e) synaptic protein levels and (f) proteostasis ? across cellular ages (DIV 7, 14 and 21). In Aim 2, we will examine the molecular mechanisms underlying heterogeneous responses of each APOE isoform to ABO-induced stress exposures. In Aim 3, we will conduct olfactory psychophysical and cognitive tasks on all participants annually to determine the influence of APOE4 status on both olfactory and cognitive trajectory over a 3-year follow-up. In view of 10% attrition rate, we will target 250 enrollees. Finally, we will validate olfactory and molecular markers of APOE4 in olfactory cells of 40 mildly cognitively impaired (MCI) and AD individuals to determine which markers predict MCI or AD status. Accomplishment of this study could result in the identification of pathophysiologic hallmarks at the earliest phases of preclinical AD, which offers a unique opportunity for effective disease modification.